【摘要】：易處理金礦的儲量正在逐年急劇減少,開采利用難處理金礦迫在眉睫。生物氧化法因其對環(huán)境友好和成本低等優(yōu)點在對難處理金礦的開采冶煉方面有廣闊的應用前景。嗜酸氧化亞鐵硫桿菌作為該法中最主要的細菌,存在著氧化能力弱、不耐砷、對各種離子不耐受等缺點,嚴重限制了生物氧化的工業(yè)化進程,所以選育高氧化活性和優(yōu)良性能菌種已成為重中之重。本論文通過利用碳離子和X射線輻照誘變嗜酸氧化亞鐵硫桿菌并選育得到具有氧化活性增高且穩(wěn)定遺傳的突變菌株,同時通過分子生物學方法檢測原始菌株和高氧化活性突變菌株的鐵氧化通路相關酶基因的表達差異,探討誘變引起該細菌氧化活性提高的分子機理。1、嗜酸氧化亞鐵硫桿菌野生型菌株的分離鑒定從陽山金礦礦坑水中純化獲得一株細菌,經(jīng)形態(tài)特征生理生化和分子鑒定表明其屬于嗜酸氧化亞鐵硫桿菌,取名YS-1。該菌長約1.2-1.5μm,寬約0.4-0.7μm,可以氧化利用二價鐵和單質硫或硫化合物作為其生長的能量來源,是典型的無機化能自養(yǎng)型微生物。菌株YS-1的最佳生長條件為:pH值2.0,搖床轉速150rpm,接種量10%,溫度35℃。值得注意的是該菌株最適生長溫度比其標準株ATCC23270高5℃,因為金礦的生物預氧化過程會散發(fā)熱量,所以菌株YS-1適合用于礦石的生物預氧化。2、碳離子和X射線誘變菌株YS-1及優(yōu)良性能菌株選育通過不同劑量的碳離子和X射線輻照對數(shù)期菌液,統(tǒng)計其致死率。結果顯示重離子輻照劑量是320Gy時,菌株的致死率達95%;在X射線輻照劑量為360Gy時致死率是82.78%。輻照之后,一方面通過耐溫和耐砷馴化分別得到可以在39℃條件下和含砷量為90mmol/L的條件下生長的突變菌株;另一方面通過選育得到兩株氧化活性提高的突變體,一株是在劑量為320Gy的碳離子輻照后篩選得到的,命名為SJ-1,其氧化速率提高非常顯著,在第48h時就可以將9K培養(yǎng)基中的二價鐵離子全部氧化,比原始菌株YS-1縮短12h。另一株是在劑量為360Gy的X射線誘變后篩選所得,命名為JW-1,其氧化活性略低于菌株SJ-1。對突變菌株SJ-1和JW-1進行傳代培養(yǎng),結果表明兩者均遺傳穩(wěn)定。3、金礦的生物預氧化和化學浸出為了檢測菌株的氧化活性對難處理金礦氧化效果的影響,將所有誘變前后的菌株用于對金礦進行生物預氧化。結果顯示突變菌株SJ-1使礦石的鐵浸出率比原始菌株YS-1增高28.12%,達到73.15%。而菌株JW-1使礦石的鐵浸出率增加到66.19%,比原始菌株提高21.16%,表明氧化活性越高的菌株對難處理金礦的作用效果越顯著。將生物氧化后的礦石用于化學浸出,結果表明經(jīng)過菌株SJ-1氧化后的礦石其金浸出率高達91.73%,比直接氰化高43.87%;而經(jīng)過菌株JW-1氧化的礦石其金浸出率達到89.89%,高于直接氰化時金浸出率42.03%。4、菌株二價鐵氧化通路相關酶基因表達的研究通過實時定量PCR技術測定突變菌株SJ-1及JW-1和原始菌株YS-1的二價鐵氧化通路相關基因在mRNA水平上的表達,發(fā)現(xiàn)碳離子和X射線輻照對菌株YS-1的鐵氧化通路相關基因表達的影響極其顯著。在突變菌株SJ-1和JW-1的各個生長階段,幾乎所有被檢測基因的表達量都高于原始菌株YS-1,且菌株SJ-1的基因上調表達更加顯著。該結果表明菌株氧化活性的提高和鐵氧化通路關鍵基因表達密切相關。
[Abstract]:The reserves of the easy-to-treat gold deposits are decreasing year by year, and it is urgent to exploit the hard-to-treat gold deposits. The biological oxidation method has wide application prospect in the mining and smelting of the refractory gold ore because of the advantages of environment-friendly and low cost and the like. Acidithiobacillus ferrooxidans, as the main bacteria in the method, has the defects of weak oxidation capacity, no arsenic, and intolerance to various ions, and severely limits the industrialization process of biological oxidation, so the breeding of high oxidation activity and excellent performance strain has become the most important. by using carbon ion and X-ray irradiation to induce the Acidithiobacillus ferrooxidans and breeding to obtain the mutant strain with increased oxidation activity and stable inheritance, at the same time, the expression difference of the related enzyme gene of the iron oxide pathway of the original strain and the high-oxidation active mutant strain is detected by a molecular biological method, the molecular mechanism of the improvement of the oxidation activity of the bacteria is explored, Isolation and identification of wild-type strain of Acidithiobacillus ferrooxidans were obtained from the pit water of Yangshan gold mine to obtain a bacterium. The morphological characteristics, physiological and biochemical characteristics and molecular identification showed that it belonged to the Acidithiobacillus ferrooxidans, named YS-1. The bacterium has a length of about 1.2 to 1.5. m u.m and a width of about 0.4 to about 0.7. m u.m, and it is possible to oxidize and utilize a divalent iron and elemental sulfur or a sulfur compound as an energy source for its growth, and is a typical autotrophic microorganism. The optimal growth conditions of strain YS-1 were: pH 2.0, shaking speed 150 rpm, inoculum 10%, temperature 35 鈩，

本文編號：2443136